Particle separation processes have a diverse variety of applications. Therapeutic techniques being developed in the medical field require efficient blood processing. Cell components of blood are typically separated by centrifugation and/or eleutriation. Improved processes for separating cell populations from blood with a minimum of undesired cell contamination are of interest to the medical field.
The following references disclose methods for separating particles from mixed populations. U.S. Pat. No. 4,350,283 discloses a centrifuge rotor adapted for continuous separation of specific particles from mixed populations thereof, by centrifugal elutriation. The rotor is adapted to be supported on a centrifuge drive shaft and has an elutriating cell housing means with at least two equally spaced apart elongated cavities disposed symmetrically with respect to the axis of rotation of the rotor. A fluid delivery spindle is disposed in the rotor having fluid inlet and outlet passages communicating with each of the elongated cavities of the rotor. An elutriating liquid is pumped through the rotor by an external pump.
G.B. Patent No. 1,313,753 disclosed a method and apparatus for conducting particle size analysis of a particulate solid material. The method comprises introducing a suspension of solid material into a rotating bowl centrifuge and causing an elutriating liquid to flow through the suspension. Elutriated particles are collected in a receiving vessel. The method is described as particularly suitable for separating particles from 0.1 to 5 microns in size from a particulate solid material. The apparatus comprises a reservoir for holding a supply of elutriating liquid, a centrifuging chamber which has an inlet and outlet and a duct means connecting the reservoir with the centrifuging chamber.
DE No. 2,918,384 (Abstract) discloses a modular system for fractionating (separating) biological cells. A cell suspension is fed into a cell disruptor and then through one or several series-connected rotors (e.g., counterflow centrifuges with elutriator/rotor). The separation of cell nuclei, particle fractions, and ribosomes is disclosed. The modular system reduces the path to be traveled by the biological material.
The following references disclose methods and apparatuses for separating blood components. U.S. Pat. No. 4,268,393 discloses an apparatus for separating platelet rich plasma (PRP) from whole blood. A blood sample is placed in a chamber and subjected to centrifugal force. A relatively small volume of saline is injected into the centrifugally outer end of the chamber, whereby PRP is displaced from the chamber into a collection bag. Red and white blood cells are held at a steady state equilibrium in the chamber. In a preferred apparatus, the saline is injected into the blood sample by driving the chamber supporting the blood sample against a saline filled collapsible cavity under the influence of centrifugal force.
U.S. Pat. No. 4,304,357 discloses an apparatus and process for separating blood into a plasma-rich component and a plasma-poor component. A processing bag containing whole blood and a flexible displacement pouch having a fluid operating diaphragm are positioned within a blood processing chamber in a centrifuge rotor. The chamber comprises a pair of contoured support shoes and a pressure plate positioned against the inner wall of the support shoe nearest the center of rotation of the rotor. As displacer fluid is introduced into the displacement pouch, blood or blood components are forced out of the processing bag and collected in a receiver container.
U.S. Pat. Nos. 4,416,654 and 4,464,167 disclose a method and apparatus for separation of specific cells. A two-port centrifugation pheresis bowl is utilized for centrifugation and high speed elutriation. In a particular embodiment, red cell-free platelets are separated from blood by pumping low density fluid, preferably plasma, through centrifugally separated cells to elutriate the cells according to their sedimentation rate.
U.S. Pat. No. 4,322,298 discloses a method and apparatus for the fractionation of a suspension of finely divided solid particles differing in sedimentation velocity. The separation of platelet rich plasma (PRP) from whole blood is accomplished by centrifuging a blood sample and injecting into its outer centrifugal end a volume of saline. The saline displaces PRP from the blood sample after red cells have been sedimented away from the other end of the bag. The preferred apparatus is designed as a closed system of interconnected bags held in a support made to fit a large centrifuge bucket.
U.S. Pat. No. 4,098,456 discloses a centrifuge cup having a mouth-defining rim, a pair of separable cap memberhalves, and means for gripping the upper end of a flexible bag position in the cup. The flexible bag is described as particularly desirable for washing blood and separating blood components. The bag contains an inlet and an outlet, plus a third, sealed port for access to blood cells after centrifugation. Blood cell washing is accomplished by percolating a wash solution through the cells.
U.S. Pat. No. 4,269,718 discloses a method and apparatus for the separation of finely divided solid particles dissimilar in size and/or density, such as platelets and other blood cells. The separation of platelets is accomplished by subjecting a blood sample to centrifugal force in a chamber and injecting a relatively small volume of saline into the centrifugally outer end of the chamber, whereby platelets are displaced from the blood sample. In the preferred apparatus, the saline is injected into the blood sample by driving the chamber supporting the blood sample into a saline filled cavity under the influence of centrifugal force.
These references disclose centrifugal elutriation processes in which particles having different sedimentation characteristics are separated from mixtures. The mixtures are subjected to a centrifugal force while a separating fluid is pumped against the force at a rate greater than the sedimentation rates of the desired particles. The separating fluid is lighter, i.e. less dense, than desired particles. Separated particles are displaced by viscous drag resulting from moving liquid on the surface of the particles. Since small-dense particles can have the same viscous drag effect as larger-less dense particles, eleutriation does not effectively separate particles having different densities. In many particle separation processes, particularly of blood cells, separation based on particle density is desirable. The process of the present invention provides improved separation of particles having different densities as compared to eleutriation.
Boyum, Scandinavian Journal of Clinical and Laboratory Investigations, 21:77 (1968) discloses the isolation of mononuclear cells and granulocytes from human blood. The reference discloses that when anticoagulated blood was layered on top of a mixture of Isopaque and Ficoll in a centrifuge tube and centrifuged, the cellular elements were divided into two main fractions: granulocytes and erythrocytes sedimented to the bottom of the tube, while mononuclear cells, together with platelets, remained at the interface. The yield of mononuclear cells was said to be almost 100 percent, when the blood was diluted with saline before centrifugation. In the process of the present invention, mononuclear cells can be separated from a mixture of blood cells in a closed apparatus and without dilution prior to separation.